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The purpose of the present study was to test the hypothesis that a preceding contractile period in isolated single skeletal muscle fibers would attenuate the decrease in pH during an identical, subsequent contractile period, thereby reducing the rate of fatigue. Intact single skeletal muscle fibers ( n = 9) were isolated from Xenopus lumbrical muscle and incubated with the fluorescent cytosolic H+ indicator 2′,7′-bis-(2-carboxyethyl)-5( 6 )-carboxyfluorescein (BCECF) AM for 30 min. Two identical contractile periods were performed in each fiber, separated by a 1-h recovery period. Force and intracellular pH (pHi) fluorescence were measured simultaneously while fibers were stimulated (tetanic contractions of 350-ms trains with 70-Hz stimuli at 9 V) at progressively increasing frequencies (0.25, 0.33, 0.5, and 1 contraction/s) until the development of fatigue (to 60% initial force). No significant difference ( P < 0.05) was observed between the first and second contractile periods in initial force development, resting pHi, or time to fatigue (5.3 ± 0.5 vs. 5.1 ± 0.6 min). However, the relative decrease in the BCECF fluorescence ratio (and therefore pHi) from rest to the fatigue time point was significantly greater ( P < 0.05) during the first contractile period (to 65 ± 4% of initial resting values) compared with the second (77 ± 4%). The results of the present study demonstrated that, when preceded by an initial fatiguing contractile period, the rise in cytosolic H+ concentration in contracting single skeletal muscle fibers during a second contractile period was significantly reduced but did not attenuate the fatigue process in the second contractile period. These results suggest that intracellular factors other than H+ accumulation contribute to the fall in force development under these conditions.

In single frog skeletal myocytes, a linear relationship exists between “fatigability” and oxidative capacity. The purpose of this investigation was to study the relationship between the intracellular Po2 (PiO2) offset kinetics and fatigability in single Xenopus laevis myocytes to test the hypothesis that PiO2 offset kinetics would be related linearly with myocyte fatigability and, by inference, oxidative capacity. Individual myocytes ( n = 30) isolated from lumbrical muscle were subjected to a 2-min bout of isometric peak tetanic contractions at either 0.25- or 0.33-Hz frequency while PiO2 was measured continuously via phosphorescence quenching techniques. The mean response time (MRT; time to 63% of the overall response) for PiO2 recovery from contracting values to resting baseline was calculated. After the initial square-wave constant-frequency contraction trial, each cell performed an incremental contraction protocol [i.e., frequency increase every 2 min from 0.167, 0.25, 0.33, 0.5, 1.0, and 2.0 Hz until peak tension fell below 50% of initial values (TTF)]. TTF values ranged from 3.39 to 10.04 min for the myocytes. The PiO2 recovery MRT ranged from 26 to 146 s. A significant ( P < 0.05), negative relationship (MRT = −12.68TTF + 168.3, r2 = 0.605) between TTF and PiO2 recovery MRT existed. These data demonstrate a significant correlation between fatigability and oxidative phosphorylation recovery kinetics consistent with the notion that oxidative capacity determines, in part, the speed with which skeletal muscle can recover energetically to alterations in metabolic demand.

The purpose of this investigation was to utilize 2,3-butanedione monoxime (BDM; an inhibitor of contractile activation) to dissociate cytosolic [Ca(2+)] ([Ca(2+)](c)) from the putative respiratory regulators that arise from muscle contraction-induced ATP utilization in order to determine the relative contribution of [Ca(2+)](c) on intracellular P(O(2)) (P(iO(2))) kinetics during the transition from rest to contractions in single skeletal myocytes isolated from Xenopus laevis lumbrical muscle. Myocytes were subjected to electrically induced isometric tetanic contractions (0.25 Hz; 2-min bouts) while peak tension and either [Ca(2+)](c) (n= 7; ratiometric fluorescence microscopy) or P(iO(2)) (n= 7; phosphorescence microscopy) was measured continuously. Cells were studied under both control and 3 mm BDM conditions in randomized order. Initial (control, 100 +/- 0%; BDM, 72.6 +/- 4.6%), midpoint (control, 86.7 +/- 1.8%; BDM, 61.6 +/- 4.1%) and end (control, 85.0 +/- 2.8%; BDM, 57.5 +/- 5.0%) peak tensions (normalized to initial control values) were significantly reduced (P0.05) with BDM compared with control (n= 14). Despite the reduced peak tension, peak [Ca(2+)](c) was not altered (P0.05) between control and BDM trials. Thus, the peak tension-to-peak [Ca(2+)](c) ratio was reduced with BDM compared with control. The absolute fall in P(iO(2)) with contractions, which is proportional to the rise in , was significantly reduced with BDM (13.2 +/- 1.3 mmHg) compared with control (22.0 +/- 2.0 mmHg). However, P(iO(2)) onset kinetics (i.e. mean response time (MRT)) was not altered between BDM (66.8 +/- 8.0 s) and control (64.9 +/- 6.3 s) trials. Therefore, the initial rate of change (defined as the fall in P(iO(2))/MRT) was significantly slower in BDM fibres compared with control. These data demonstrate in these isolated single skeletal muscle fibres that unchanged peak [Ca(2+)](c) in the face of reduced metabolic feedback from the contractile sites evoked with BDM did not alter P(iO(2)) onset kinetics in isolated single frog myocytes, suggesting that metabolic signals arising from the contractile sites play a more substantial role than [Ca(2+)](c) in the signalling pathway to oxidative phosphorylation during the transition from rest to repeated tetanic contractions.

During the past decade, unenhanced computed tomography (CT) has become the standard of reference in the detection of urinary calculi owing to its high sensitivity (>95%) and specificity (>98%) in this setting. Numerous diseases may manifest as acute flank pain and mimic urolithiasis. Up to one-third of unenhanced CT examinations performed because of flank pain may reveal unsuspected findings unrelated to stone disease, many of which can help explain the patient's condition. Alternative diagnoses are most commonly related to gynecologic conditions (especially adnexal masses) and nonstone genitourinary disease (eg, pyelonephritis, renal neoplasm), closely followed by gastrointestinal disease (especially appendicitis and diverticulitis). Hepatobiliary, vascular, and musculoskeletal conditions may also be encountered. Vascular causes of acute flank pain must always be considered, since these constitute life-threatening emergencies that may require the intravenous administration of contrast material for diagnosis. Radiologists must be familiar with the typical findings of urinary stone disease at unenhanced CT, as well as the spectrum of alternative diagnoses that may be detected with this modality, to accurately diagnose the source of flank pain.

This study investigated the effects of acute creatine kinase (CK) inhibition (CKi) on contractile performance, cytosolic Ca2+concentration ([Ca2+]c), and intracellular Po2(Pi[Formula: see text]) in Xenopus laevis isolated myocytes during a 2-min bout of isometric tetanic contractions (0.33-Hz frequency). Peak tension was similar between trials during the first contraction but was significantly ( P < 0.05) attenuated for all subsequent contractions in CKivs. control (Con). The fall in Pi[Formula: see text](ΔPi[Formula: see text]) from resting values was significantly greater in Con (26.0 ± 2.2 Torr) compared with CKi(17.8 ± 1.8 Torr). However, the ratios of Con to CKiend-peak tension (1.53 ± 0.11) and ΔPi[Formula: see text](1.49 ± 0.11) were similar, suggesting an unaltered aerobic cost of contractions. Additionally, the mean response time (MRT) of ΔPi[Formula: see text]was significantly faster in CKivs. Con during both the onset (31.8 ± 5.5 vs. 49.3 ± 5.7 s; P < 0.05) and cessation (21.2 ± 4.1 vs. 68.0 ± 3.2 s; P < 0.001) of contractions. These data demonstrate that initial phosphocreatine hydrolysis in single skeletal muscle fibers is crucial for maintenance of sarcoplasmic reticulum Ca2+release and peak tension during a bout of repetitive tetanic contractions. Furthermore, as Pi[Formula: see text]fell more rapidly at contraction onset in CKicompared with Con, these data suggest that CK activity temporally buffers the initial ATP-to-ADP concentration ratio at the transition to an augmented energetic demand, thereby slowing the initial mitochondrial activation by mitigating the energetic control signal (i.e., ADP concentration, phosphorylation potential, etc.) between sites of ATP supply and demand.

The purpose of the present study was to compare the individual fatigue characteristics of isolated single skeletal muscle fibres with their mitochondrial volume density (MVD), using direct histological morphometry. Single muscle fibres (n= 14) were microdissected from lumbrical muscle of adult female Xenopus laevis, and force was measured while fibres were stimulated (tetanic contractions of 200 ms trains with 70 Hz stimuli at 9 V) at progressively increasing frequencies (2 min each at 0.25, 0.33, 0.5 and 1 contractions s(-1)) until fatigue (50% initial maximal force) had been established. Following the end of the fatigue protocol, MVD was determined by electron microscopy. Time to fatigue varied among the individual fibres from 3.3 to 10 min. MVD of individual fibres ranged from 3.0 to 9.2% and was positively correlated (r= 0.93) with time to fatigue of corresponding fibres. These results, using direct histological measurements of MVD: (1) support on a single cell basis the notion that oxidative capacity is a major determinant of muscle fatigue resistance; and (2) show that the fatigue profile of a single cell can be used to predict oxidative capacity.

We tested the hypotheses that: (1) Ca 2 + handling and force production would be irreversibly altered in skeletal muscle during steady-state contractions when subjected to severe, prolonged hypoxia and subsequent reoxygenation; and (2) application of the cardio-protective drug trimetazidine would attenuate these alterations. Single, living skeletal muscle fibres from Xenopus laevis were injected with the Ca 2 + indicator fura 2, and incubated for 1 h prior to stimulation in 100 μM TMZ-Ringer solution (TMZ; n = 6) or standard Ringer solution (CON; n = 6). Force and relative free cytosolic Ca 2 + concentration ([Ca 2 + ] c ) were measured during continuous tetanic contractions produced every 5 s as fibres were sequentially perfused in the following manner: 3 min high extracellular P o 2 (159 mmHg), 15 min hypoxic perfusion (3-5 mmHg) then 3 min high P o 2 . Hypoxia caused a decrease in force and peak [Ca 2 + ] c in both the TMZ and CON fibres, with no significant (P < 0.05) difference between groups. However, basal [Ca 2 + ] c was significantly lower during hypoxia in the TMZ group vs. the CON group. While reoxygenation generated only modest recovery of relative force and peak [Ca 2 + ] c in both groups, basal [Ca 2 + ] c remained significantly less in the TMZ group. These results demonstrated that in contracting, single skeletal muscle fibres, TMZ prevented increases in basal [Ca 2 + ] c generated during a severe hypoxic insult and subsequent reoxygenation, yet failed to protect the cell from the deleterious effects of prolonged hypoxia followed by reoxygenation.

The lumped parameter describing skeletal muscle diffusional conductance for O(2), DM(O(2)), reflects all of the resistances for O(2) in moving from red cell to muscle fiber mitochondria. The purpose of our study was to determine if the carotenoid compound, trans sodium crocetinate (TSC), which has been reported to increase the diffusivity of O(2) in plasma, improves DM(O(2)) and thus, V(O(2),max) in maximally contracting in situ skeletal muscle. V(O(2),max) was measured in the isolated perfused canine gastrocnemius (n=5) during 3 min of isometric tetanic contractions at 1 Hz, while the animal was breathing 12% O(2) (PA(O(2))=32+/-2 Torr, mean+/-S.E.) under two experimental conditions. The first was a control contraction period and the second (following 60 min recovery) was performed within 5 min after infusion of a 0.1 mg x ml(-1) solution of TSC (total dose 100 microg kg(-1)). There were no significant differences in convective O(2) delivery (11.9+/-2.3 vs. 12.1+/-2.2 ml x min(-1) x 100 g(-1)), V(O(2),max) (9.5+/-1.5 vs. 9.6+/-1.5 ml x min(-1) x 100 g(-1)) or calculated DM(O(2)) (0.37+/-0.03 vs. 0.37+/-0.04 ml x min(-1) x 100 g(-1) x Torr(-1)) between contraction periods. As such, our results show that TSC does not improve performance in maximally contracting canine gastrocnemius muscle in situ under moderately hypoxic conditions, suggesting either that TSC in this situation does not increase plasma O(2) diffusivity or that this step in O(2) diffusion from red cell to myocyte does not constrain DM(O(2)).

Reperfusion following prolonged ischemia induces cellular damage in whole skeletal muscle models. Ischemic preconditioning attenuates the deleterious effects. We tested whether individual skeletal muscle fibers would be similarly affected by severe hypoxia and reoxygenation (H/R) in the absence of extracellular factors and whether cellular damage could be alleviated by hypoxic preconditioning. Force and free cytosolic Ca2+([Ca2+]c) were monitored in Xenopus single muscle fibers ( n = 24) contracting tetanically at 0.2 Hz during 5 min of severe hypoxia and 5 min of reoxygenation. Twelve cells were preconditioned by a shorter bout of H/R 1 h before the experimental trial. In preconditioned cells, force relative to initial maximal values (P/Po) and relative peak [Ca2+]cfell ( P< 0.05) during 5 min of hypoxia and recovered during reoxygenation. In contrast, P/Poand relative peak [Ca2+]cfell more during hypoxia ( P < 0.05) and recovered less during reoxygenation ( P < 0.05) in control cells. The ratio of force to [Ca2+]cwas significantly higher in the preconditioned cells during severe hypoxia, suggesting that changes in [Ca2+]cwere not solely responsible for the loss in force. We conclude that 1) isolated skeletal muscle fibers contracting in the absence of extracellular factors are susceptible to H/R injury associated with changes in Ca2+handling; and 2) hypoxic preconditioning improves contractility, Ca2+handling, and cell recovery during subsequent hypoxic insult.

Although evidence for muscle O2 diffusion limitation of maximal O2 uptake has been found in the intact organism and isolated muscle, its relationship to diffusion distance has not been examined. Thus we studied six sets of three purpose-bred littermate dogs (aged 10–12 mo), with 1 dog per litter allocated to each of three groups: control (C), exercise trained for 8 wk (T), or left leg immobilized for 3 wk (I). The left gastrocnemius muscle from each animal was surgically isolated, pump-perfused, and electrically stimulated to peak O2 uptake at three randomly applied levels of arterial oxygenation [normoxia, arterial[Formula: see text]([Formula: see text]) 77 ± 2 (SE) Torr; moderate hypoxia, [Formula: see text]: 33 ± 1 Torr; and severe hypoxia, [Formula: see text]: 22 ± 1 Torr]. O2 delivery (ml ⋅ min−1 ⋅ 100 g− 1) was kept constant among groups for each level of oxygenation, with O2 delivery decreasing with decreasing [Formula: see text]. O2extraction (%) was lower in I than T or C for each condition, but calculated muscle O2 diffusing capacity (Dm[Formula: see text]) per 100 grams of muscle was not different among groups. After the experiment, the muscle was perfusion fixed in situ, and a sample from the midbelly was processed for microscopy. Immobilized muscle showed a 45% reduction of muscle fiber cross-sectional area ( P < 0.05), and a resulting 59% increase in capillary density ( P < 0.05) but minimal reduction in capillary-to-fiber ratio (not significant). In contrast, capillarity was not significantly different in T vs. C muscle. The results show that a dramatically increased capillary density (and reduced diffusion distance) after short-term immobilization does not improve Dm[Formula: see text] in heavily working skeletal muscle.